Identification and regulation of a gene required for cell fusion during mating of the yeast Saccharomyces cerevisiae.

Abstract

We have devised a screen for genes from the yeast Saccharomyces cerevisiae whose expression is affected by cell type or by the mating pheromones. From this screen we identified a gene, FUSI, whose pattern of expression revealed interesting regulatory strategies and whose product was required for efficient cell fusion during mating. Transcription of FUSI occurred only in a and a cells, not in a/a cells, where it was repressed by al -a2, a regulatory activity present uniquely in ala cells. Transcription of FUSJ showed an absolute requirement for the products of five STE genes, STE4, STES, STE7, STEII, and STE12. Since the activators STE4, STE5, and STE12 are themselves repressed by al a2, the failure to express FUSI in a/a cells is probably the result of a cascade of regulatory activities; repression of the activators by al a2 in turn precludes transcription of FUSI. In addition to regulation of FUSI by cell type, transcription from the locus increased 10-fold or more when a or a cells were exposed to the opposing mating pheromone. To investigate the function of the Fusl protein, we created fusi null mutants. In fusi x fusl matings, the cells of a mating pair adhered tightly and appeared to form zygotes. However, the zygotes were abnormal. Within the conjugation bridge they contained a partition that prevented nuclear fusion and mixing of organelles. The predicted sequence of the Fusl protein (deduced from the FUSI DNA sequence) and subcellular fractionation studies with Fus1-pgalactosidase hybrid proteins suggest that Fusl is a membrane or secreted protein. Thus, Fusl may be located at a position within the cell where it is poised to catalyze cell wall or plasma membrane fusion.The yeast Saccharomyces cerevisiae exhibits three distinct cellular phenotypes: a, a, and a/a. The a and a cell types, which are typically haploid, can mate to yield the third cell type, an a/a diploid. These a/a diploids are not capable of mating but can be induced to undergo meiosis and sporulation, thereby regenerating haploid a and a cells (reviewed in references 18, 19, and 41). To mate efficiently, a and a cells must each secrete a specific peptide pheromone to which only the other cell type can respond. Thus, a cells secrete a 13-amino-acid-residue peptide, a factor, which binds to a specific receptor on the surface of a cells and triggers a physiological response in those cells (20,21). Similarly, a cells display a surface receptor that enables them to respond to the a-factor pheromone secreted by a cells (3,15,31). The response of a and a cells to the pheromone of the other is similar and includes an increase in the transcription of a small set of genes (16,17,43), an increase in the ability to agglutinate with the other cell type (4, 13), and arrest of the cell division cycle in the Gl phase (8,48). A mating pair whose cell cycles have been synchronized in this fashion can undergo controlled cell wall dissolution and membrane fusion to yield an a/a zygote.Despite the complexity of the cellular phenotypes outlined above, the three...

Coupling of cell identity to signal response in yeast: interaction between the alpha 1 and STE12 proteins.

“…We also compared the ability of the two homologs to mediate transcriptional activation in S. cerevisiae on three different cell type-specific genes: MFA2, an a-specific gene encoding a-factor (Michaelis and Herskowitz 1988), MFal, an a-specific gene encoding or-factor (Kurjan and Herskowitz 1982;Flessel et al 1989), and FUS1, a haploid-specific gene necessary for cell fusion (McCaffrey et al 1987;Trueheart et al 1987). In each case the upstream regulatory region of the cell type-specific gene was linked to the E. coli lacZ gene.…”

“…First, cell identity itself is functionally equivalent to the synthesis of the particular set of pheromone and receptor that allows a cell to be responsive to the opposite cell type (see Bender and Sprague 1989). Second, the transcription of the a-and o~-specific genes, as well as of certain haploid-specific genes, is dependent on an intact pheromone response pathway (Hartwell 1980;Fields and Herskowitz 1985;Hartig et al 1986;McCaffrey et al 1987;Fields et al 1988;Hagen et al 1991). Third, in general, transcription of only cell type-specific genes is inducible by pheromone treatment.…”

Dual Lipid Modification Motifs in Gαand GγSubunits Are Required for Full Activity of the Pheromone Response Pathway inSaccharomyces cerevisiae

“…Cells in early logarithmic phase (A 600 , 0.35-0.5) were treated with various concentrations of ␣-factor for 2 h. ␤-Galactosidase activity in permeabilized cells was determined (McCaffrey et al, 1987) Gpa1p protein levels were determined by immunoblot of 100 g of yeast total cell lysate from CMY23 and CMY24. Endogenous Gpa1p was detected with 9126, a polyclonal antibody against Gpa1p (generously provided by D. Stone, University of Illinois, Chicago, IL).…”

CDC36 and CDC39 are negative elements in the signal transduction pathway of yeast.

“…(McCaffrey et al, 1987). The FUS 1 gene cannot be induced in strains that carry a ste mutation (McCaffrey et al, 1987). In contrast, strains that are defective in SCG 1 express FUS 1 constitutively (Nakayama et al, 1988).…”

“…These studies allow us to place CDC36 and CDC39 in a specific position in the signaling pathway. (McCaffrey et al, 1987). The FUS 1 gene cannot be induced in strains that carry a ste mutation (McCaffrey et al, 1987).…”